RESUMO
Autosomal dominant hereditary spastic paraplegia (AD-HSP) is a genetically heterogeneous neurodegenerative disorder characterized by progressive spasticity of the lower limbs. Among the four loci causing AD-HSP identified so far, the SPG4 locus at chromosome 2p2-1p22 has been shown to account for 40-50% of all AD-HSP families. Using a positional cloning strategy based on obtaining sequence of the entire SPG4 interval, we identified a candidate gene encoding a new member of the AAA protein family, which we named spastin. Sequence analysis of this gene in seven SPG4-linked pedigrees revealed several DNA modifications, including missense, nonsense and splice-site mutations. Both SPG4 and its mouse orthologue were shown to be expressed early and ubiquitously in fetal and adult tissues. The sequence homologies and putative subcellular localization of spastin suggest that this ATPase is involved in the assembly or function of nuclear protein complexes.
Assuntos
Adenosina Trifosfatases/genética , Mutação , Paraplegia Espástica Hereditária/genética , Adenosina Trifosfatases/química , Adenosina Trifosfatases/metabolismo , Motivos de Aminoácidos , Sequência de Aminoácidos , Animais , Sequência de Bases , Células Cultivadas , Clonagem Molecular , Análise Mutacional de DNA , Éxons/genética , Etiquetas de Sequências Expressas , Humanos , Íntrons/genética , Camundongos , Mitocôndrias Musculares/metabolismo , Dados de Sequência Molecular , Fosforilação Oxidativa , RNA Mensageiro/análise , RNA Mensageiro/genética , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Paraplegia Espástica Hereditária/enzimologia , Paraplegia Espástica Hereditária/metabolismo , Paraplegia Espástica Hereditária/patologia , EspastinaRESUMO
The approximately 10 kbp region encompassing nprB and argJ at 102 degrees on the Bacillus subtilis chromosome was sequenced, revealing 12 ORFs, four known genes (argJ, argC, ipi and nprB) and two genes, yitY and yitS, whose products respectively display significant homology with L-gulono-gamma-lactone oxidase of rat and dihydrofolate reductase of Staphylococcus aureus. The data also indicated that nprB mapped to a different position than previously published.
Assuntos
Bacillus subtilis/genética , Cromossomos Bacterianos/genética , DNA Bacteriano/genética , Genes Bacterianos , Fases de Leitura Aberta , L-Gulonolactona Oxidase , Dados de Sequência Molecular , Mapeamento por Restrição , Análise de Sequência de DNA , Especificidade da Espécie , Desidrogenase do Álcool de Açúcar/genética , Tetra-Hidrofolato Desidrogenase/genéticaRESUMO
Autosomal dominant hereditary spastic paraplegia (AD-HSP) is a genetically heterogeneous disorder characterized by progressive spasticity of the lower limbs. A major locus (SPG4) causing AD-HSP in about 40% of the families was mapped to chromosome 2p. The analysis of six SPG4-linked AD-HSP families using the RED procedure previously showed the expansion of a CAG repeat in affected individuals. To identify the gene responsible for this form of HSP, we have constructed a 3.5-Mb YAC contig flanked by loci D2S400 and D2S367, have subcloned five of these YACs spanning the candidate region into cosmids, and screened these cosmid libraries for the presence of CAG repeat sequences. Four CAG repeats have been identified but none of them is expanded in 26 patients from 13 SPG4-linked AD-HSP families. A gene map comprising 21 transcripts was established using expressed sequence tags (ESTs) assigned previously to this region of 2p21-p22 with radiation hybrid panels GeneBridge 4 and G3. Full-length cDNAs corresponding to the 14 ESTs mapping to the SPG4 interval flanked by loci D2S352 and D2S2347 were isolated and sequenced. None contains a CAG repeat in its coding sequence. Finally, we have assembled a BAC contig composed of 37 clones that were also screened for the presence of CAG repeats; this failed to detect additional repeats to those identified on YACs.
Assuntos
Cromossomos Humanos Par 2/genética , Paraplegia Espástica Hereditária/genética , Repetições de Trinucleotídeos/genética , Cromossomos Bacterianos/genética , Clonagem Molecular , Mapeamento de Sequências Contíguas , Etiquetas de Sequências Expressas , Humanos , Repetições de Microssatélites , Análise de Sequência de DNARESUMO
Autosomal dominant familial spastic paraplegia (AD-FSP) is a genetically heterogeneous neurodegenerative disorder characterized by progressive spasticity of the lower limbs. Three loci on chromosome 14q (SPG3), 2p (SPG4), and 15q (SPG6) were shown to be responsible for AD-FSP. Analysis of recombination events in three SPG3-linked families allowed us to narrow the critical interval from 9 to 5 cM. An approximately 5-Mb YAC contig comprising 32 clones and 90 STSs was built from D14S301 to D14S991, encompassing this region of 14q21. Fifty-six ESTs assigned previously to this region with radiation hybrid (RH) panels Genebridge 4 and G3 were precisely localized on the YAC contig. The 90 STSs positioned on the contig were tested on the TNG RH panel to compare our YAC-based map with an RH map at a high level of resolution. Comparison between our map and the whole genome mapping data on this interval of chromosome 14q is discussed.
Assuntos
Cromossomos Humanos Par 14/genética , Genoma Humano , Paraplegia Espástica Hereditária/genética , Mapeamento Cromossômico , Mapeamento de Sequências Contíguas , Etiquetas de Sequências Expressas , Saúde da Família , Feminino , Humanos , Células Híbridas/efeitos da radiação , Masculino , Repetições de Microssatélites , Linhagem , Sitios de Sequências Rotuladas , Transcrição GênicaRESUMO
Autosomal dominant hereditary spastic paraplegia (AD-HSP) is a group of genetically heterogeneous neurodegenerative disorders characterized by pro- gressive spasticity of the lower limbs. Five AD-HSP loci have been mapped to chromosomes 14q, 2p, 15q, 8q and 12q. The SPG4 locus at 2p21-p22 has been shown to account for approximately 40% of all AD-HSP families. SPG4 encoding spastin, a putative nuclear AAA protein, has recently been identified. Here, sequence analysis of the 17 exons of SPG4 in 87 unrelated AD-HSP patients has resulted in the detection of 34 novel mutations. These SPG4 mutations are scattered along the coding region of the gene and include all types of DNA modification including missense (28%), nonsense (15%) and splice site point (26.5%) mutations as well as deletions (23%) and insertions (7.5%). The clinical analysis of the 238 mutation carriers revealed a high proportion of both asymptomatic carriers (14/238) and patients unaware of symptoms (45/238), and permitted the redefinition of this frequent form of AD-HSP.
Assuntos
Adenosina Trifosfatases/genética , Genes Dominantes , Mutação , Paraplegia/genética , Adenosina Trifosfatases/fisiologia , Adolescente , Adulto , Idoso , Criança , Códon sem Sentido , Genótipo , Humanos , Pessoa de Meia-Idade , Dados de Sequência Molecular , Mutação de Sentido Incorreto , Fenótipo , Polimorfismo Genético , Splicing de RNA , EspastinaRESUMO
We report the construction of a tiling path of around 650 clones covering more than 99% of human chromosome 14. Clone overlap information to assemble the map was derived by comparing fully sequenced clones with a database of clone end sequences (sequence tag connector strategy). We selected homogeneously distributed seed points using an auxiliary high-resolution radiation hybrid map comprising 1,895 distinct positions. The high long-range continuity and low redundancy of the tiling path indicates that the sequence tag connector approach compares favourably with alternative mapping strategies.